α-, β-, and γ-Cyclodextrins, which consist of 6, 7, and 8 glucose molecules that are linked each other via the α-1,4 glucosyl linkage, respectively, have been known as cyclic saccharides composed of glucose units. These cyclodextrins have been used in a variety of fields due to their advantageous inherent properties of non-reducibility, tasteless, enclosing hydrophobic materials, etc. There have been being pursued remarkable researches directed to improve properties of cyclodextrins and impart additional new functions thereupon. For example, Japanese Patent Kokai Nos. 9,708/94, 14,789/94, 16,705/94, 298,806/94, and 25,305/98 proposed branched cyclodextrins with different branching structures, which are prepared by coupling a glycosyl group such as a glucosyl, galactosyl, mannosyl, glucosaminyl, or N-acetylglucosaminyl group to cyclodextrins; processes for producing the same; and uses thereof.
As an example of cyclic saccharide reported recently, there is a cyclic tetrasaccharide, reported by Gregory L. Cote et al. in European Journal of Biochemistry, Vol. 226, pp. 641-648 (1994), composed of glucose molecules linked each other via the alternating α-1,3 and α-1,6 bonds and having the structures represented by Chemical Formulae A and B as bonding fashions between atoms and between glucosyl groups, respectively. In addition to Chemical Formulae A and B, cyclo{→6)-α-D-glucopyranosyl-(1→3)-α-D-glucopyranosyl-(1→6)-α-D-glucopyranosyl-(1-3)-α-D-glucopyranosyl-(1→} belongs to the above cyclic tetrasaccharide. Throughout the specification, the term “cyclotetrasaccharide” means the above-identified cyclic tetrasaccharide.
Chemical Formula B:cyclo{→6)-α-D-Glcp-(1→3)-α-D-Glcp-(1→6)-α-D-Glcp-(1→3)-α-D-Glcp-(1→}
The above report by Cote et al. shows that cyclotetrasaccharide is formed by allowing alternan, a type of polysaccharide composed of glucose molecules linked via the alternating α-1,3 and α-1,6 bonds to act on alternanase, a type of hydrolyzing enzyme. Thereafter, cyclotetrasaccharide has been expected to be used in different fields similarly as in or much more useful than conventional cyclodextrins. The method of the report, however, may not suitable for an industrial-scale production of cyclotetrasaccharide, because alternan used as a starting material is not easily obtainable and the yield of cyclotetrasaccharide from the material is insufficient in view of industrial-scale production.
The same applicant as the present invention disclosed an α-isomaltosyl-transferring enzyme, a novel enzyme which forms cyclotetrasaccharide when acts on a saccharide having a glucose polymerization degree of at least three and having both an isomaltosyl group at the non-reducing end and the α-1,4 glucosyl bond as a linkage other than the linkage at the non-reducing end (abbreviated as “α-isomaltosylglucosaccharide” hereinafter) as disclosed in Japanese Patent Application No. 149,484/2000, and Japanese Patent Application No 229,557/2000 (International Publication No. WO 01/90,338 A1) applied for based on the above Japanese Patent Application; and also disclosed an α-isomaltosylglucosaccharide-forming enzyme, a novel enzyme which forms α-isomaltosylglucosaccharide when acts on a maltooligosaccharide having a glucose polymerization degree of at least three, as disclosed in Japanese Patent Application No. 233,364/2000 and Japanese Patent Application No. 234,937/2000 (International Publication No. WO 02/10,361 A1), applied for based on Japanese Patent Application No.233,364/2000. Further, the above applicant proposed a method to produce cyclotetrasaccharide as a main product from starch, a widely, commonly used material for producing foods by using the α-isomaltosylglucosaccharide-forming enzyme and the α-isomaltosyl-transferring enzyme in combination, as disclosed in Japanese Patent Application Nos. 233,364/2000 and 234,937/2000 (International Publication No. WO 02/10,361 A1). This proposal was a breakthrough for an industrial scale production of cyclotetrasaccharide.
Thus, the study on cyclotetrasaccharide has just merely been started, and further studies for elucidating unknown functions and developing new uses of cyclotetrasaccharide are now being greatly expected. Even though cyclotetrasaccharide is a known compound, there is found no study to produce derivatives thereof as a main object, because cyclotetrasaccharide has not yet been easily obtained. So far found is merely the above report by Cote et al. that reported only a 6-O-glucopyranosyl derivative of cyclotetrasaccharide, represented by Chemical Formula C, isolated and identified as a by-product in a negligible yield through the action of alternanase on alternan. Chemical Formula D represents the 6-O-glucopyranosyl derivative in terms of bonding fashions between glucosyl groups.


Similarly as in cyclodextrins, supplying of glycosyl derivatives of cyclotetrasaccharide would provide a useful knowledge for developing uses of cyclotetrasaccharide through analyses on their properties, and also it remarkably influences on the development of uses of novel saccharides, obtained by improving or modifying the properties and functions of cyclotetrasaccharide.